Nguyen, Canh Vinh Ngoc (2021) Investigating Cancer Cell Sensitivity to Anticancer Drugs in Relation to Heat Shock Factor 1 Expression. Research Master thesis, Victoria University.

Abstract

Breast cancer (BrCa) is the most common cancer diagnosed in women, with many patients progressing to advanced stages where prognosis is poor and morbidity more likely. Ninety per cent of patients will succumb to their disease primarily due to metastasis. Metastatic cancer cells are invasive, migratory and highly resistant to standard chemotherapies. Therefore, greater knowledge is needed to characterize mechanisms by which cancer cells become resistant to anticancer agents. A mechanism by which cancer cells may develop resistance is through stimulating stress response pathways such as the heat shock response (HSR). This pathway is regulated by heat shock factor 1 (HSF1), which transcriptionally regulates transcription of heat shock proteins (HSP) as well as many non-HSPs. HSPs protect normal cells during exposure to proteotoxic stresses; however, HSF1 has also been found to facilitate pro-metastatic pathways in cancer, distinct from the canonical HSR. HSF1 expression is known to be significantly increased in multiple cancers and significantly correlates with poor clinical outcomes; yet little is known regarding the role of HSF1 in the resistance/sensitivity of cancer cells towards anticancer therapeutics. To address this, the aim of this project was to examine whether HSF1 may have a direct role in mediating anticancer drug sensitivity in cancer cells and whether anticancer drugs stimulate HSF1 activation. To achieve this, a series of doxycycline-inducible HSF1 knockdown (KD) BrCa cell lines, T47D and MDA-MB-231, were generated. From previous bioinformatic studies, HSF1 was identified as potentially mediating the sensitivity of cancer cells to several anticancer drugs. A number of these drugs were screened in both the T47D and MDA-MB-231 series of doxycycline-inducible HSF1 KD cells. From these screens, it was identified that loss of HSF1 resulted in a significant decrease in the sensitivity of MDA-MB-231 cells towards the EGFR inhibitor, Lapatinib, but this was not evident in the less advanced T47D BrCa cells. However, the T47D cells were found to be increased in their sensitivity to doxorubicin with HSF1 knockdown. To determine whether anticancer drugs stimulated the HSR, T47D and MDA-MB-231 heat shock element (HSE) bioluminescent reporter cells were generated. Cells were successfully generated to express firefly luciferase under the control of HSE, indicative of HSF1 activity and the quantitative assessment of anticancer drug induced- stress. However, these reporter cell models revealed the previously undetermined impact of drug vehicles (DMSO, EtOH) upon HSE activation indicative of the potential for false positives within drug screens if not properly controlled. This work has identified that HSF1 plays a role in mediating the sensitivity of aggressive BrCa cells to the EGFR inhibitor, Lapatinib. Conversely, HSF1 mediates resistance to doxorubicin in the less aggressive T47D BrCa cells. Moreover, the use of HSE reporter cells to determine HSR activation by anticancer drugs needs stringent controls in relation to drug vehicles due to the potential for these vehicles to activate the HSR leading to false positives within anticancer drug screens.

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